The renal proximal tubule (PT) reabsorbs ~80% of the HCO-3 filtered at the glomerulus. Previous work on isolated perfused rabbit and mouse PTs shows that the rate of HCO-3 reabsorption (JHCO3) markedly rises with increases in basolateral [CO2] ([CO2]BL) or decreases in [HCO-3]BL, but is not altered by changes in pHBL. Thus, the PT must have a mechanism for sensing CO2/HCO-3. The CO2-evoked increase in JHCO3 is blocked by antagonists of apical ANG II AT1a receptors and by specific inhibitors of the ErbB family of receptor tyrosine kinases (e.g., ErbB1). We were intrigued to learn of an orphaned receptor protein tyrosine phosphatase (RPTP3) with a putative extracellular ligand binding domains that strongly resembles carbonic anhydrase (CA). Preliminary data suggest that knocking out RPTP3 eliminates the JHCO3 response to both ?[CO2]BL and ?[HCO-3]BL. The three aims are a multidisciplinary approach to address, at three levels of integration, how the PT senses ?[CO2]BL and alters JHCO3: (1) Perfused PTs. What is the role of RPTP3? Do elevations in [CO2]BL enhance the response to added luminal ANG II or ANG-(1-7)? Does chronic acidosis enhance the CO2-evoked increase in JHCO3? And are AT1a-null and RPTP3-null mice more sensitive to chronic acidosis? (2) Biochemistry in PT suspensions. Does CO2/HCO-3 produce a unique RPTP3-dependent phosphotyrosine fingerprint on ErbB1? Can a proteomic approach identify downstream - targets of CO2/RPTP3/ErbB1? (3) The RPTP3 molecule. Is it CO2 or HCO3 that activates the phosphatase activity of RPTP3? Where is RPTP3 located? Can mutations in the CA- like domain of RPTP3 generate CA activity? Can a CA inhibitor bind to the CA-like domain? What is the crystal structure of the CA-like domain? The proposed work will illuminate a highly novel mechanism for sensing CO2 and or HCO-3-independent of changes in pH-and could have important implications for clinical approaches to acidosis, hypertension, and cancer. PUBLIC HEALTH RELEVANCE: The acid-base status of the body is predominantly maintained by secretion of bodily H+ (acid) across the proximal tubule (PT) cells of the kidney into fluid that is eventually excreted as urine. Acid secretion by PT cells is not directly stimulated by increased body acid per se-but by the ratio of body CO2/HCO-3 (the components of soda water), which itself is acid-sensitive. The goals of this project are to identify the CO2-sensing protein in the PT and its downstream targets, and to understand the highly novel mechanism by which increased CO2 increases acid secretion. The proposed work could have important implications for clinical approaches to acidosis, hypertension, and cancer.